Upon completion of an Internal Medicine residency at the Medical College of Wisconsin, Timothy W. Behrens, M.D. served a year as Chief Medical Resident before embarking on a three year combined clinical and research Fellowship in Rheumatology at Milwaukee. With the support of the Arthritis Foundation and the Leukemia Society of America, he then took a three year research fellowship in the laboratory of Dr. Louis M. Staudt at the National Cancer Institute - NIH applying the tools of modern molecular biology to the study of the immune system. His major project at NIH was the cloning of a novel lymphoid-restricted gene called Jawl, a developmentally regulated protein localized to the endoplasmic reticulum of lymphocytes. As a Junior faculty member at the University of Minnesota, Dr. Behrens has worked closely with his proposed sponsor, Dr. Brian Van Ness, Professor of Biochemistry and Human Genetics, to initiate a new line of investigation characterizing the role of bcl-x, a gene related to the bcl-2 oncogene, in the regulation of apoptosis in the immune system. In the last year, Dr. Behrens cloned cDNAs for the mouse bcl-x gene, identified a novel transmembrane deleted splice variant that is functional in blocking apoptosis, and showed that bcl-x is highly inducible in lymphocytes. Recent studies demonstrate that overexpressed bcl - x protects the immature B lymphocyte cell line WEHI 231 from the apoptosis triggered by crosslinking of surface immunoglobulin M (IgM), oxidant stress, serum deprivation, and gamma-irradiation. The proposed experiments will further characterize the regulation of mbcl-x protein and mRNA expression in this B cell line and will study the signals that control expression in normal mouse B lymphocytes. Using a transgenic mouse model, the in vivo influence of overexpressed bcl-x on B lineage development and function will be examined. The mechanism by which bcl-x blocks apoptosis will also be examined with an emphasis on whether bcl-x participates in an intracellular antioxidant pathway. Finally, proteins that interact with blc-x will be identified, and dominant negative approaches to disruption of bcl-x function will be explored. The long term goals of the project are to further our understanding of late events in the apoptotic pathway and to develop strategies for interrupting the function of bcl-2 family members for therapeutic benefit in human disease.